大片形吸虫病早期诊断夹心ELISA方法的建立及应用  

Development and Application of Sandwich ELISA for Diagnosis of Fascioliasis giganticain Early Stage

作　　者：李雪[1] 魏志鹏[1] 赵明龙[1] 许心婷 刘宇[1] 吴文德[1] 蒋薇[1] 黄甜[1] 龙飞翔 陈汉忠[1] 

机构地区：[1]广西大学动物科学与技术学院,南宁530004

出　　处：《中国畜牧兽医》2016年第4期899-905,共7页China Animal Husbandry & Veterinary Medicine

基　　金：广西研究生教育创新计划项目(YCSZ2014026)

摘　　要：为建立一种早期诊断大片形吸虫病的试验方法,本试验对5株分泌抗大片形吸虫分泌排泄抗原(ES)单克隆抗体的杂交瘤细胞进行复苏、制备单克隆抗体,配对筛选出7D1、7D2单克隆抗体,将7D2作为包被抗体,7D1作为酶标抗体,通过条件优化,建立早期诊断大片形吸虫病的夹心ELISA方法。结果显示,当包被抗体稀释度为1∶6 400(0.208μg/mL)、采用5%脱脂奶粉封闭、酶标记抗体稀释度为1∶10 000(0.200μg/mL)时,最早可检测到感染此病第7天小鼠的血清循环抗原,其敏感性可达到1∶3 200(0.156μg/mL),与其他几种寄生虫抗原均无交叉反应,具有较高的特异性和稳定性。本试验成功建立了早期检测大片形吸虫病的夹心ELISA方法,为大片形吸虫病的诊治及开发早期诊断大片形吸虫病的试剂盒奠定了基础,具有临床应用价值。To establish a method for the diagnosis of Fascioliasis giganticain early stage,five hybridoma cell lines were recovered and used for preparation of monoclonal antibodies.7D2 was used as a capture antibody and 7D1 as detection antibody.Coating antibody dilution was 1∶6 400（0.208μg/mL）,5% nonfat milk was used as blocking solution and detection antibody dilution was 1∶10 000（0.200μg/mL）.The detection limit of the sandwich ELISA was 1∶3 200（0.156μg/mL）,with good specificity and stability,and there was no cross reaction with other kinds of parasite antigen.The results showed that the method provided the important conditions and theoretical basis for the diagnosis of Fascioliasis gigantica in early stage.This would save unnecessary economic losses to the livestock,and it had clinical application value.

关 键 词：早期诊断 单克隆抗体 大片形吸虫 夹心ELISA 

分 类 号：R446.61[医药卫生—诊断学]